High-throughput screening based identification of small molecule antagonists of integrin CD11b/CD18 ligand binding

Binding of leukocyte specific integrin CD11b/CD18 to its physiologic ligands is important for the development of normal immune response in vivo. Integrin CD11b/CD18 is also a key cellular effector of various inflammatory and autoimmune diseases. However, small molecules selectively inhibiting the function of integrin CD11b/CD18 are currently lacking. We used a newly described cell-based high-throughput screening assay to identify a number of highly potent antagonists of integrin CD11b/CD18 from chemical libraries containing >100,000 unique compounds. Computational analyses suggest that the identified compounds cluster into several different chemical classes. A number of the newly identified compounds blocked adhesion of wild-type mouse neutrophils to CD11b/CD18 ligand fibrinogen. Mapping the most active compounds against chemical fingerprints of known antagonists of related integrin CD11a/CD18 shows little structural similarity, suggesting that the newly identified compounds are novel and unique.     

Efficiency clustering for low-density microarrays and its application to QPCR

Background  

Pathway-targeted or low-density arrays are used more and more frequently in biomedical research, particularly those arrays that are based on quantitative real-time PCR. Typical QPCR arrays contain 96-1024 primer pairs or probes, and they bring with it the promise of being able to reliably measure differences in target levels without the need to establish absolute standard curves for each and every target. To achieve reliable quantification all primer pairs or array probes must perform with the same efficiency.     

Comparative analysis of the zeta-crystallin/quinone reductase gene in guinea pig and mouse

zeta-Crystallin is a novel nicotinamide adenine dinucleotide phosphate:quinone reductase, present at enzymatic levels in various tissues of different species, which is highly expressed in the lens of some hystricomorph rodents and camelids. We report here the complementary DNA (cDNA) cloning of zeta-crystallin from liver libraries in guinea pig (Cavia porcellus), where zeta-crystallin is highly expressed in the lens, and in the laboratory mouse (Mus musculus), where expression in the lens occurs only at enzymatic levels. A 5' untranslated sequence different from the one previously reported for the guinea pig lens cDNA was found in these clones. We also report the isolation of genomic clones including the complete guinea pig zeta-crystallin gene and the 5' region of this gene in mouse. These results show the presence of two promoters in the guinea pig zeta-crystallin gene, one responsible for expression at enzymatic levels and the other responsible for the high expression in the lens. The guinea pig lens promoter is not present in the mouse gene. This is the first example in which the recruitment of an enzyme as a lens crystallin can be explained by the acquisition of an alternative lens- specific promoter.      

Small molecule agonists of integrin CD11b/CD18 do not induce global conformational changes and are significantly better than activating antibodies in reducing vascular injury

Background

CD11b/CD18 is a key adhesion receptor that mediates leukocyte adhesion, migration and immune functions. We recently identified novel compounds, leukadherins, that allosterically enhance CD11b/CD18-dependent cell adhesion and reduce inflammation in vivo, suggesting integrin activation to be a novel mechanism of action for the development of anti-inflammatory therapeutics. Since a number of well-characterized anti-CD11b/CD18 activating antibodies are currently available, we wondered if such biological agonists could also become therapeutic leads following this mechanism of action.

Methods

We compared the two types of agonists using in vitro cell adhesion and wound-healing assays and using animal model systems. We also studied effects of the two types of agonists on outside-in signaling in treated cells.

Results

Both types of agonists similarly enhanced integrin-mediated cell adhesion and decreased cell migration. However, unlike leukadherins, the activating antibodies produced significant CD11b/CD18 macro clustering and induced phosphorylation of key proteins involved in outside-in signaling. Studies using conformation reporter antibodies showed that leukadherins did not induce global conformational changes in CD11b/CD18 explaining the reason behind their lack of ligand-mimetic outside-in signaling. In vivo, leukadherins reduced vascular injury in a dose-dependent fashion, but, surprisingly, the anti-CD11b activating antibody ED7 was ineffective.

Conclusions

Our results suggest that small molecule allosteric agonists of CD11b/CD18 have clear advantages over the biologic activating antibodies and provide a mechanistic basis for the difference.

General significance

CD11b/CD18 activation represents a novel strategy for reducing inflammatory injury. Our study establishes small molecule leukadherins as preferred agonists over activating antibodies for future development as novel anti-inflammatory therapeutics.     

Ovarian activity and plasma sex steroid levels of Distichodus antonii in relation to environmental conditions in the upper basin of the Congo River

Distichodus antonii is an endemic fish species of the Congo River basin in which the stocks of wild populations are threatened by overfishing pressure. Knowledge of its reproductive biology would be useful in consideration of conservation and management options for the species. Therefore, this study investigated changes in ovarian activity and levels of steroid profiles in wild populations in relation to variation in temperature and rainfall. Adult females (n = 101, body weight of 3 183 ± 14.75 g, SE) were captured monthly over one year (2013–2014). Apart from evaluation of oocyte diameters and gonad developmental stages, gonado-, hepato-, lipososomatic indices (GSI, HSI, LSI) and plasma levels of sex steroids (testosterone-T, estradiol-17β-E2) were determined. The results suggested a synchronous development of oocytes with two annual reproductive seasons over the one-year study. Plasma T and E2 levels peaked during spawning periods likely reflecting active oogenesis. The highest values of morphosomatic indices were observed during the longest rainfall period in September, and were associated with high steroidogenic activity evidenced by increased E2 production. In addition, more vitellogenic oocytes (September and October) were observed during the latter season than during the short rainy season (in May).     

The development of pistillate and perfect florets in Xeranthemum squarrosum (Asteraceae)

The formation of capitulum inflorescence with two different types of floret is an interesting issue in floral biology and evolution. Here we studied the inflorescence, floral ontogeny and development of the everlasting herb, Xeranthemum squarrosum, using epi‐illumination microscopy. The small vegetative apex enlarged and produced involucral bracts with helical phyllotaxy, which subtended floret primordia in the innermost whorl. Initiation of floret primordia was followed by an acropetal sequence, except for pistillate peripheral florets. The origin of receptacular bracts was unusual, as they derived from the floral primordia rather than the receptacular surface. The order of whorl initiation in both disc and pistillate flowers included corolla, androecium and finally calyx, together with the gynoecium. The inception of sepals and stamens occurred in unidirectional order starting from the abaxial side, whereas petals incepted unidirectionally from the adaxial or abaxial side. Substantial differences were observed in flower structure and the development between pistillate and perfect florets. Pistillate florets presented a zygomorphic floral primordium, tetramerous corolla and androecium and two sepal lobes. In these florets, two sepal lobes and four stamen primordia stopped growing, and the ovary developed neither an ovule nor a typical stigma. The results suggest that peripheral pistillate florets in X. squarrosum, which has a bilabiate corolla, could be considered as an intermediate state between ancestral bilabiate florets and the derived ray florets.     

AIR: A batch-oriented web program package for construction of supermatrices ready for phylogenomic analyses

Background  

Large multigene sequence alignments have over recent years been increasingly employed for phylogenomic reconstruction of the eukaryote tree of life. Such supermatrices of sequence data are preferred over single gene alignments as they contain vastly more information about ancient sequence characteristics, and are thus more suitable for resolving deeply diverging relationships. However, as alignments are expanded, increasingly numbers of sites with misleading phylogenetic information are also added. Therefore, a major goal in phylogenomic analyses is to maximize the ratio of information to noise; this can be achieved by the reduction of fast evolving sites.     

Gallic acid-based indanone derivatives as anticancer agents

Gallic acid-based indanone derivatives have been synthesised. Some of the indanones showed very good anticancer activity in MTT assay. Compounds 10, 11, 12 and 14 possessed potent anticancer activity against various human cancer cell lines. The most potent indanone (10, IC₅₀ = 2.2 μM), against MCF-7, that is, hormone-dependent breast cancer cell line, showed no toxicity to human erythrocytes even at higher concentrations (100 μg/ml, 258 μM). While, indanones 11, 12 and 14 showed toxicities to erythrocytes at higher concentrations.