Binding immunoglobulin protein resolves rheumatoid synovitis: a xenogeneic study using rheumatoid arthritis synovial membrane transplants in SCID mice

Introduction  

Binding immunoglobulin protein (BiP) has previously shown powerful anti-inflammatory properties in the collagen-induced arthritis (CIA) model, where a single dose of BiP has proved to be both a long-term prophylactic and therapeutic. In both CIA and human in vitro studies, BiP induced regulatory T cells. The present investigation looked at the anti-inflammatory effect of BiP on inflamed human synovial tissue transplanted into severe combined immunodeficient mice (SCID), a chimaeric in vivo model previously used to test the efficacy of biologic therapies.     

Functional polymorphisms in the TERT promoter are associated with risk of serous epithelial ovarian and breast cancers

Genetic variation at the TERT-CLPTM1L locus at 5p15.33 is associated with susceptibility to several cancers, including epithelial ovarian cancer (EOC). We have carried out fine-mapping of this region in EOC which implicates an association with a single nucleotide polymorphism (SNP) within the TERT promoter. We demonstrate that the minor alleles at rs2736109, and at an additional TERT promoter SNP, rs2736108, are associated with decreased breast cancer risk, and that the combination of both SNPs substantially reduces TERT promoter activity.     

Expression profile of two storage-protein gene families in hexaploid wheat revealed by large-scale analysis of expressed sequence tags

To discern expression patterns of individual storage-protein genes in hexaploid wheat (Triticum aestivum cv Chinese Spring), we analyzed comprehensive expressed sequence tags (ESTs) of common wheat using a bioinformatics technique. The gene families for alpha/beta-gliadins and low molecular-weight glutenin subunit were selected from the EST database. The alignment of these genes enabled us to trace the single nucleotide polymorphism sites among both genes. The combinations of single nucleotide polymorphisms allowed us to assign haplotypes into their homoeologous chromosomes by allele-specific PCR. Phylogenetic analysis of these genes showed that both storage-protein gene families rapidly diverged after differentiation of the three genomes (A, B, and D). Expression patterns of these genes were estimated based on the frequencies of ESTs. The storage-protein genes were expressed only during seed development stages. The alpha/beta-gliadin genes exhibited two distinct expression patterns during the course of seed maturation: early expression and late expression. Although the early expression genes among the alpha/beta-gliadin and low molecular-weight glutenin subunit genes showed similar expression patterns, and both genes from the D genome were preferentially expressed rather than those from the A or B genome, substantial expression of two early expression genes from the A genome was observed. The phylogenetic relationships of the genes and their expression patterns were not correlated. These lines of evidence suggest that expression of the two storage-protein genes is independently regulated, and that the alpha/beta-gliadin genes possess novel regulation systems in addition to the prolamin box.     

Factors influencing the sporulation and cyst formation of Aphanomyces invadans, etiological agent of ulcerative mycosis in Atlantic menhaden, Brevoortia tyrannus

Oomycete infections caused by Aphanomyces invadans occur in freshwater and estuarine fishes around the world. Along the east coast of the USA, skin ulcers caused by A. invadans are prevalent in Atlantic menhaden, Brevoortia tyrannus. From laboratory observations low salinities appear crucial to transmission of the pathogen. To better understand aspects of transmission, we characterized sporulation and cyst formation of secondary zoospores of two isolates of A. invadans at different salinities and temperatures. Sporulation occurred only at low salinities. At room temperature (ca. 20-22 C), using "pond water" augmented with artificial sea salts, the endemic strain WIC and the Thailand strain PA7 of A. invadans produced free-swimming secondary zoospores at salinities of 0, 1 and 2 psu (practical salinity unit = per thousand), but not at 4 psu or higher. Secondary zoospores of another species, ATCC-62427 (Aphanomyces sp.), were observed at 1, 2, 4 and 8 psu but not at 0 and 12 psu. Secondary zoospores of all three isolates, especially WIC, were abundant and motile 1-2 d postsporulation. Sporulation was temperature dependent and occurred over a relatively narrow range. No sporulation occurred at 4, 30 or 35 C for either WIC or PA7. For both strains zoospore production within 1-3 d after the initiation of sporulation was more prolific at 25 C than at 20 and 15 C. At 15 C production of zoospores was sustained over 11 d for WIC and 5 d for PA7. At room temperature single WIC secondary zoospores remained motile 12-18 h. Salinities exceeding 4 psu or vigorous shaking caused immediate cyst formation of WIC secondary zoospores. Exposure to menhaden tissue, but not tissues of other fishes to secondary zoospores (WIC), caused rapid (2 h) cyst formation. Cysts were capable of excysting when transferred to 1 psu water within 2-3 h of cyst formation. Cysts that had remained encysted in 6.5 psu for 24 h did not excyst when transferred to 1 psu water. Salinity and temperature requirements for sporulation indicate that juvenile menhaden must acquire infections during rain or in low salinity oligohaline waters.     

Polymorphisms of TNFalpha, IL1beta, and IL1RN genes in chronic obstructive pulmonary disease

It is recognized that genetic factors play a role in the susceptibility to COPD. COPD is characterized by airflow limitation. Chronic inflammation causes small airway disease and parenchymal destruction, leading to the airflow limitation. Polymorphisms in pro-inflammatory cytokine genes may confer a risk for the development of COPD. A case-control association study was performed in Japanese population (88 COPD patients and 61 controls) and Egyptian population (106 patients and 72 controls). Genotype and allele frequencies of the TNFalpha -308 G/A and +489 G/A polymorphisms, the IL1beta -511 C/T, -31 T/C, and +3954 C/T polymorphisms, and a VNTR polymorphism in intron 2 of the IL1RN gene were investigated. In addition, pairwise haplotype frequencies were analyzed. When studied independently, none of the polymorphisms were associated with the development of COPD in both populations. However, in the Egyptian population, the distributions of the haplotype (IL1beta -31 T/C : IL1beta +3954 C/T) were significantly different between the COPD patients and the controls (p(corr)=0.0037). Our findings suggest that this haplotype within the IL1beta gene may be involved in the pathogenesis of COPD and that the genetic factors of COPD susceptibility might be different between different populations.     

An Arabidopsis aspartic protease functions as an anti-cell-death component in reproduction and embryogenesis

The components and pathways that regulate and execute developmental cell death programmes in plants remain largely unknown. We have found that the PROMOTION OF CELL SURVIVAL 1 (PCS1) gene in Arabidopsis, which encodes an aspartic protease, has an important role in determining the fate of cells in embryonic development and in reproduction processes. The loss-of-function mutation of PCS1 causes degeneration of both male and female gametophytes and excessive cell death of developing embryos. Conversely, ectopic expression of PCS1 causes the septum and stomium cells that normally die in the anther wall to survive instead, leading to a failure in anther dehiscence and male sterility. PCS1 provides a new avenue for understanding the mechanisms of the programmed cell death processes that are associated with developmental pathways in plants and makes available a useful tool for engineering the male sterility trait for hybrid seed production.     

Characterization of the organic cation transporter SLC22A16: a doxorubicin importer

Specific efflux transporters, such as P-glycoprotein, have been shown to confer drug resistance by decreasing the intracellular accumulation of anticancer drugs. Understanding influx transporters, as well as efflux transporters, is essential to overcome this resistance. We report the expression profile and pharmacological characterization of an organic cation transporter, SLC22A16. The results of our experiments indicate that SLC22A16 is a mediator of doxorubicin uptake in cancer cells. Quantitative real-time RT-PCR analyses show that SLC22A16 is expressed in primary samples taken from patients with acute leukemia. Xenopus oocytes injected with SLC22A16 cRNA import doxorubicin, a widely used anticancer drug for hematological malignancies, in a saturable and dose-dependent manner. The apparent Km value for doxorubicin import was 5.2+/-0.4 microM. In cytotoxic assays, stable transfectants of leukemic Jurkat cells overexpressing SLC22A16 cells became significantly more sensitive to doxorubicin (2 microM) treatment. Characterization of SLC22A16 will help in designing novel therapies targeting hematological malignancies.     

Kinetic studies of DNA cleavage reactions catalyzed by an ATP-dependent deoxyribonuclease on a 27-MHz quartz-crystal microbalance

Catalytic DNA cleavage reactions by an ATP-dependent deoxyribonuclease (DNase) from Micrococcus luteus were monitored directly with a DNA-immobilized 27-MHz quartz-crystal microbalance (QCM). The 27-MHz QCM is a very sensitive mass-measuring device in aqueous solution, as the frequency decreases linearly with increasing mass on the electrode at a nanogram level. Three steps in ATP-dependent DNA hydrolysis reactions, including (1) binding of DNase to the end of double-stranded DNA (dsDNA) on the QCM electrode (mass increase), (2) degradation of one strand of dsDNA in the 3' --> 5' direction depending on ATP (mass decrease), and (3) release of the enzyme from the nonhydrolyzed 5'-free-ssDNA (mass decrease), could be monitored stepwise from the time dependencies of QCM frequency changes. Kinetic parameters for each step were obtained as follows. The binding constant (K(a)) of DNase to the dsDNA was determined as (28 +/- 2) x 10(6) M(-)(1) (k(on) = (8.0 +/- 0.3) x 10(3) M (-)(1) s(-)(1) and k(off) = (0.29 +/-0.01) x 10(-)(3) s(-)(1)), and it decreased to (0.79 +/- 0.16) x 10(6) M(-)(1) (k'(on) = (2.3 +/- 0.2) x 10(3) M (-)(1) s(-)(1) and k'(off) = (2.9 +/- 0.1) x 10(-)(3) s(-)(1)) for the completely nonhydrolyzed 5'-free ssDNA. This is the reason the DNase bound to the dsDNA substrate can easily release from the nonhydrolyzed 5'-free-ssDNA after the complete hydrolysis of the 3' --> 5' direction of the complementary ssDNA. K(a) values depended on the DNA structures on the QCM, and the order of these values was as follows: the dsDNA having a 4-base-mismatched base-pair end (3) > the dsDNA having a 5' 15-base overhanging end (2) > the dsDNA having a blunt end (1) > the ssDNA having a 3'-free end (4) > the ssDNA having a 5'-free end (5). Thus, DNase hardly recognized the free 5' end of ssDNA. Michaelis-Menten parameters (K(m) for ATP and k(cat)) of the hydrolysis process also could be obtained, and the order of k(cat)/K(m) was as follows: the dsDNA having a blunt end (1) approximately the dsDNA having a 4-base-mismatched base-pair end (3) > the ssDNA having a free 3' end (4) > the ssDNA having a free 5' end (5). Thus, DNase could not recognize and not hydrolyze the free 5' end of ssDNA. The DNA hydrolysis reaction could be driven by dATP and GTP (purine base) as well as ATP, whereas the cleavage efficiency was very low driven with UTP, CTP (pyrimidine base), ADP, and AMP.