The integument of the paddlefish,Polyodon spathula

The integument of the paddlefish (Polyodon spathula) is unusual as a relatively small amount of mucus is produced by epithelial cells that are not modified into regular mucous gland cells. A thick compact epidermis and dermis compensate for the slight amount of mucus secreted. Paddlefish have a variety of scales formed of concentric bony lamellae containing osteocytes. There are five kinds of scales: dorsal and ventral fulcra on the caudal fin, rhomboidal scales on the caudal lobe, horny denticles over the pectoral girdle, calcareous denticles on the trunk, and anchor-shaped plates on the rostrum. Except for the fulcra, the scales are undoubtedly vestigial. The numerous surface pits on the rostrum, head, operculum, and throat are epithelial invaginations which are not connected to lateral line canals. No nerves lead to the pits. The spherical to cuboidal and often ciliated cells at the base of the pits are considered to be aplasic cells of unformed neuromasts.     

Computer modeling of fibrin polymerization kinetics correlated with electron microscope and turbidity observations: clot structure and assembly are kinetically controlled.

Although much is known about fibrin polymerization, because it is complex, the effects of various modifications are not intuitively obvious and many experimental observations remain unexplained. A kinetic model presented here that is based on information about mechanisms of assembly accounts for most experimental observations and allows hypotheses about the effects of various factors to be tested. Differential equations describing the kinetics of polymerization were written and then solved numerically. The results have been related to turbidity profiles and electron microscope observations. The concentrations of intermediates in fibrin polymerization, and fiber diameters, fiber and protofibril lengths have been calculated from these models. The simplest model considered has three steps; fibrinopeptide A cleavage, protofibril formation, and lateral aggregation of protofibrils to form fibers. The average number of protofibrils per fiber, which is directly related to turbidity, can be calculated and plotted as a function of time. The lag period observed in turbidity profiles cannot be accurately simulated by such a model, but can be simulated by modifying the model such that oligomers must reach a minimum length before they aggregate. Many observations, reported here and elsewhere, can be accounted for by this model; the basic model may be modified to account for other experimental observations. Modeling predicts effects of changes in the rate of fibrinopeptide cleavage consistent with electron microscope and turbidity observations. Changes only in the rate constants for initiation of fiber growth or for addition of protofibrils to fibers are sufficient to account for a wide variety of other observations, e.g., the effects of ionic strength or fibrinopeptide B removal or thrombospondin. The effects of lateral aggregation of fibers has also been modeled: such behavior has been observed in turbidity curves and electron micrographs of clots formed in the presence of platelet factor 4. Thus, many aspects of clot structure and factors that influence structure are directly related to the rates of these steps of polymerization, even though these effects are often not obvious. Thus, to a large extent, clot structure is kinetically determined.     

Crystals of modified fibrinogen: Size,shape and packing of molecules

We have produced several new macroscopic crystal forms and a variety of microcrystals from modified flbrinogens. Bovine and rabbit flbrinogens crystallize after limited digestion by a bacterial protease or α-chymotrypsin. The fibrinogens making up these crystals are largely intact and highly clottable. Tentative molecular packing arrangements for two crystal forms have been deduced. The crystal morphology and cleavage planes were used in this analysis. The characteristic α-helical coiled-coil reflections arid spikes of intensity in certain directions in the crystal X-ray patterns serve as markers for the orientation of the fibrinogen molecules. Changes that occur in one of the forms during preparation for electron microscopy, as shown by comparison with X-ray experiments on crystals in various stains and solvents, support this packing model. These studies provide preliminary evidence that fibrinogen is about 450 Å in length and that the molecules bond end-to-end to form filaments making up the crystals.     

Versatile Solid Supports for Oligonucleotide Synthesis that Incorporate Urea Bridge

The universal solid support, USIII, representing a new and improved version of commercial USII, as well as 2 ′-deoxynucleoside and 2 ′-deoxy-2 ′-fluoronucleoside bound supports, incorporating a labile phenoxyacetyl fragment, was synthesized by an aminomethyl polystyrene carbamoylation with corresponding azides in the presence of aqueous triethylammonium bicarbonate. All three solid phases incorporate a stable urea tether, thus bridging the polymer and functional linker. These new matrices proved to be potent solid phases for the synthesis of DNA, RNA, or modified oligonucleotides as well as randomized mixed 2 ′-ribo/2 ′-deoxy-2 ′-fluoro-RNA libraries and/or DNA libraries, randomized with trinucleotides (codons).     

Long‐term repopulation of aged bone marrow stem cells using young Sca‐1 cells promotes aged heart rejuvenation

Reduced quantity and quality of stem cells in aged individuals hinders cardiac repair and regeneration after injury. We used young bone marrow (BM) stem cell antigen 1 (Sca‐1) cells to reconstitute aged BM and rejuvenate the aged heart, and examined the underlying molecular mechanisms. BM Sca‐1⁺ or Sca‐1^? cells from young (2–3 months) or aged (18–19 months) GFP transgenic mice were transplanted into lethally irradiated aged mice to generate 4 groups of chimeras: young Sca‐1⁺, young Sca‐1^?, old Sca‐1⁺, and old Sca‐1^?. Four months later, expression of rejuvenation‐related genes (Bmi1, Cbx8, PNUTS, Sirt1, Sirt2, Sirt6) and proteins (CDK2, CDK4) was increased along with telomerase activity and telomerase‐related protein (DNA‐PKcs, TRF‐2) expression, whereas expression of senescence‐related genes (p16^INK4a, P19^ARF, p27^Kip1) and proteins (p16^INK4a, p27^Kip1) was decreased in Sca‐1⁺ chimeric hearts, especially in the young group. Host cardiac endothelial cells (GFP^?CD31⁺) but not cardiomyocytes were the primary cell type rejuvenated by young Sca‐1⁺ cells as shown by improved proliferation, migration, and tubular formation abilities. C‐X‐C chemokine CXCL12 was the factor most highly expressed in homed donor BM (GFP⁺) cells isolated from young Sca‐1⁺ chimeric hearts. Protein expression of Cxcr4, phospho‐Akt, and phospho‐FoxO3a in endothelial cells derived from the aged chimeric heart was increased, especially in the young Sca‐1⁺ group. Reconstitution of aged BM with young Sca‐1⁺ cells resulted in effective homing of functional stem cells in the aged heart. These young, regenerative stem cells promoted aged heart rejuvenation through activation of the Cxcl12/Cxcr4 pathway of cardiac endothelial cells.     

The IGF1 small dog haplotype is derived from Middle Eastern grey wolves: a closer look at statistics,sampling, and the alleged Middle Eastern origin of small dogs

This paper is a response to Gray MM, Sutter NB, Ostrander EA, Wayne RK: The IGF1 small dog haplotype is derived from Middle Eastern grey wolves. BMC Biology 2010, 8:16.     

The molecular structure of human tissue type XV presents a unique conformation among the collagens

Establishing the structure of the non-fibrillar collagens has provided a unique perspective to understanding their specialized functions in the extracellular matrix. These proteins exhibit very diverse conformations and supramolecular assemblies. Type XV collagen is a large macromolecule distinguished by a highly interrupted collagenous domain and many utilized sites of attachment for CS (chondroitin sulfate) and HS (heparan sulfate) glycosaminoglycan chains. It is present in most basement membrane zones of human tissues, where it is found closely associated with large collagen fibrils. To determine the molecular shape and organization of type XV, the protein was purified from human umbilical cords by salt extraction, and by ion-exchange and antibody-affinity chromatography. The representation of type XV in one of its most abundant tissue sources is estimated at only (1-2)x10(-4)% of dry weight. The molecules examined by transmission electron microscopy after rotary shadowing were visualized in multiple forms. Relatively few type XV monomers appeared elongated and kinked; most molecules were found in a knot/figure-of-eight/pretzel configuration not previously described for a collagen. Collective measurements of these populations revealed an average length of 193+/-16 nm. At the N-terminal end, identified by C-terminal antibody binding, were three 7.7 nm-diameter spheres, corresponding to TSPN-1 (N-terminal module of thrombospondin-1) modules, and attached to the collagen backbone by a short linker. The type XV monomers show the ability to self-assemble into higher-order structures. Some were arranged in complex clusters, but simpler oligomers, which may represent intermediates, were observed in a cruciform pattern with intermolecular binding sites that probably originate in the interruption sequences. The morphology of type XV is thus the antithesis of the fibrillar collagens, and the shape attains the required flexibility to form the spectrum of interconnecting links between banded fibrils at the basement membrane/interstitial border. These type XV structures may act as a biological 'spring' to stabilize and enhance resilience to compressive and expansive forces, and the multimers, in particular, with selective complements of many localized CS and HS chains, may be instrumental in spatial and temporal recruitment of modulators in growth, development and pathological processes.     

An overview of malaria transmission from the perspective of Amazon Anopheles vectors

In the Americas, areas with a high risk of malaria transmission are mainly located in the Amazon Forest, which extends across nine countries. One keystone step to understanding the Plasmodium life cycle in Anopheles species from the Amazon Region is to obtain experimentally infected mosquito vectors. Several attempts to colonise Ano- pheles species have been conducted, but with only short-lived success or no success at all. In this review, we review the literature on malaria transmission from the perspective of its Amazon vectors. Currently, it is possible to develop experimental Plasmodium vivax infection of the colonised and field-captured vectors in laboratories located close to Amazonian endemic areas. We are also reviewing studies related to the immune response to P. vivax infection of Anopheles aquasalis, a coastal mosquito species. Finally, we discuss the importance of the modulation of Plasmodium infection by the vector microbiota and also consider the anopheline genomes. The establishment of experimental mosquito infections with Plasmodium falciparum, Plasmodium yoelii and Plasmodium berghei parasites that could provide interesting models for studying malaria in the Amazonian scenario is important. Understanding the molecular mechanisms involved in the development of the parasites in New World vectors is crucial in order to better determine the interaction process and vectorial competence.     

An IGF-I promoter polymorphism modifies the relationships between birth weight and risk factors for cardiovascular disease and diabetes at age 36

BACKGROUND: Biochemical testing for pheochromocytoma by measurement of fractionated plasma metanephrines is limited by false positive rates of up to 18% in people without known genetic predisposition to the disease. The plasma normetanephrine fraction is responsible for most false positives and plasma normetanephrine increases with age. The objective of this study was to determine if we could improve the specificity of fractionated plasma measurements, by statistically adjusting for age. METHODS: An age-adjusted metanephrine score was derived using logistic regression from 343 subjects (including 33 people with pheochromocytoma) who underwent fractionated plasma metanephrine measurements as part of investigations for suspected pheochromocytoma at Mayo Clinic Rochester (derivation set). The performance of the age-adjusted score was validated in a dataset of 158 subjects (including patients 23 with pheochromocytoma) that underwent measurements of fractionated plasma metanephrines at Mayo Clinic the following year (validation dataset). None of the participants in the validation dataset had known genetic predisposition to pheochromocytoma. RESULTS: The sensitivity of the age-adjusted metanephrine score was the same as that of traditional interpretation of fractionated plasma metanephrine measurements, yielding a sensitivity of 100% (23/23, 95% confidence interval [CI] 85.7%, 100%). However, the false positive rate with traditional interpretation of fractionated plasma metanephrine measurements was 16.3% (22/135, 95% CI, 11.0%, 23.4%) and that of the age-adjusted score was significantly lower at 3.0% (4/135, 95% CI, 1.2%, 7.4%) (p < 0.001 using McNemar's test). CONCLUSION: An adjustment for age in the interpretation of results of fractionated plasma metanephrines may significantly decrease false positives when using this test to exclude sporadic pheochromocytoma. Such improvements in false positive rate may result in savings of expenditures related to confirmatory imaging.     

The disintegrin echistatin stabilizes integrin alphaIIbbeta3's open conformation and promotes its oligomerization

We have employed echistatin, a 5.4 kDa snake venom disintegrin, as a model protein to investigate the paradox that small ligand-mimetics can bind to the resting alphaIIbbeta3 integrin while adhesive macromolecules cannot. We characterized the interactions between purified human alphaIIbbeta3 and two recombinant echistatin variants: rEch (1-49) M28L, chosen for its selectivity toward beta3-integrins, and rEch (1-40) M28L, a carboxy-terminal truncation mutant. While both contain an RGD integrin targeting sequence, only rEch (1-49) M28L was an effective inhibitor of alphaIIbbeta3 function. Electron microscopy of rotary shadowed specimens yielded a variety of alphaIIbbeta3 conformers ranging from compact, spherical particles (maximum dimension 22 nm) to the classical "head with two tails" forms (32 nm). The population of larger particles (42-56 nm) increased from 17% to 28% in the presence of rEch (1-49) M28L, indicative of ligand-induced oligomerization. Sedimentation velocity measurements demonstrated that both full length and truncated echistatin perturbed alphaIIbbeta3's solution structure, yielding slower-sedimenting open conformers. Dynamic light scattering showed that rEch (1-49) M28L protected alphaIIbbeta3 from thermal aggregation, raising its transition mid-point from 46 degrees C to 69 degrees C; a smaller shift resulted with rEch (1-40) M28L. Sedimentation equilibrium demonstrated that both echistatin ligands induced substantial alphaIIbbeta3 dimerization. van't Hoff analysis revealed a pattern of entropy/enthalpy compensation similar to tirofiban, a small RGD ligand-mimetic that binds tightly to alphaIIbbeta3, but yields smaller conformational perturbations than echistatin. We propose that echistatin may serve as a paradigm for understanding multidomain adhesive macromolecules because its ability to modulate alphaIIbbeta3's structure resides on an RGD loop, while full disintegrin activity requires an auxiliary site that includes the carboxy-terminal nine amino acid residues.